Effects of dietary lectins on ion transport in epithelia

Abstract

Phytohemagglutinins are widely distributed in common food items. They constitute a heterogeneous group of proteins, which are often resistant to proteolysis in the gastrointestinal tract. Upon binding to the luminal membrane of intestinal cells, they can interfere with digestive, protective or secretory functions of the intestine. Phytohemagglutinins present in red kidney beans and jackbeans have been shown to induce diarrhea and hypersecretion in human airways, but the underlying mechanisms remain obscure. We examined how agglutinins from wheat germ (WGA), soy bean (SBA), red kidney beans (Pha-E, Pha-L), and jackbeans (Con-A) affect ion transport in mouse airways and large intestine using Ussing chamber techniques. We found that Pha-E, Pha-L, and Con-A but not WGA and SBA inhibit electrogenic Na(+) absorption dose dependently in both colon and trachea. The inhibitory effects of Con-A on Na(+) absorption were suppressed by the sugar mannose, by inhibition of phospholipase C (PLC) and protein kinase C (PKC). Thus, nutritional phytohemagglutinins block salt absorption in a PLC- and PKC-dependent manner, probably by inhibition of the epithelial Na(+) channel (ENaC). This effect may be therapeutically useful in patients suffering from cystic fibrosis.

Figure 1

(a) Original recording of the transepithelial voltage V_te in a mouse colonic epithelium. The voltage deflections (ΔV_te) are induced by pulsed current injection (0.5 μA). Effects of amiloride (A, 10 μmol l⁻¹) before and after exposure (20 min) to concanavalin-A (100 μg ml⁻¹). (b) Summary of the effects of the lectins concanavalin-A (Con-A), erythroagglutinin (Pha-E), leuagglutinin (Pha-L), soy bean agglutinin (SBA), and wheat germ agglutinin (WGA) (all 100 μgm l⁻¹) on amiloride-sensitive Na⁺ absorption (I_sc−Amil) in mouse colonic epithelia, when applied to either luminal or basolateral sides. (c) Summary of the short circuit currents I_sc induced by Con-A (100 μmol l⁻¹), carbachol (100 μmol l⁻¹), and IBMX (100 μmol l⁻¹) together with forskolin (10 μmol l⁻¹) in mouse colon in the presence of amiloride. (d) Summary of the effects of amiloride (10 μmol l⁻¹) applied under control conditions, after replacing extracellular Na⁺ by N-methyl D-glucamine (NMDG), and after addition of NMDG in the presence of NMDG. ^* indicate significant difference when compared to control (number of experiments).

Figure 2

(a) Original recording of the transepithelial voltage V_te in a tracheal epithelium. Effects of amiloride (A, 10 μmol l⁻¹) before and after exposure (20 min) to concanavalin-A (100 μg ml⁻¹). (b) Summary of the effects of the lectins concanavalin-A (Con-A), erythroagglutinin (Pha-E), leuagglutinin (Pha-L), soy bean agglutinin (SBA), and wheat germ agglutinin (WGA) (all 100 μg ml⁻¹) on amiloride-sensitive Na⁺ absorption (I_sc−Amil) in mouse colonic epithelia, when applied to either luminal or basolateral sides. (c–e) Dose–response of the effects of the lectins Con-A, Pha-E, and Pha-L on ion transport (I_sc) in mouse trachea. ^* indicate significant difference when compared to control (number of experiments).

Figure 3

(a) Original recording of the transepithelial voltage V_te in mouse trachea. At 20 min incubation of increasing concentrations of Con-A reduced V_te and the inhibitory effect of amiloride on V_te. (b) Incubation for the same time period in control ringer solution did not reduce V_te and did not attenuate amiloride responses. (c) Dose–response curve for the inhibitory effects of Con-A on I_sc measured in tracheas of homozygous G551D-CFTR mice. (d) Concentration dependence of the inhibitory effect of amiloride on I_sc detected in mouse trachea in the absence or presence of Con-A (number of experiments).

Figure 4

Effects of inhibitors of signal transduction on lectin effects in mouse trachea. (a) Summary of the effects on I_sc of luminal application of mannose (10 mM) and Con-A (100 μmol l⁻¹) in the presence of mannose. (b) Summary of the effects on I_sc of luminal/basolateral application of the PLC inhibitor U73122 (10 μM) and Con-A (100 μmol l⁻¹) in the presence of U73122. (c–h) Effects of Ca²⁺ chelating by 30 min preincubation with BAPTA-AM (10 μmol l⁻¹) and inhibiting protein kinase C by 30 min preincubation with BIM (1 μmol l⁻¹) on lectin induced changes in transport. (c, e, g) Subsequent application of amiloride (10 μmol l⁻¹) under control conditions, after incubation in BAPTA, and after addition of the lectin in the presence of BAPTA. (d, f, h) Subsequent application of amiloride (10 μmol l⁻¹) under control conditions, after incubation in BIM, and after addition of the lectin in the presence of BIM. ^* indicate significant difference compared to control (paired t-test) (number of experiments).